Azathioprine response

Genetic polymorphisms influence TPMT and NUDT15 activity. Several published studies indicate that patients with reduced TPMT or NUDT15 activity receiving usual doses of 6-MP or azathioprine, accumulate excessive cellular concentrations of active 6-TGNs, and are at higher risk for severe myelosuppression. Because of the risk of toxicity, patients with TPMT or NUDT15 deficiency require alternative therapy or dose modification.

Approximately 0.3% (1:300) of patients of European or African ancestry have two loss-of-function alleles of the TPMT gene and have little or no TPMT activity (homozygous deficient or poor metabolizers), and approximately 10% of patients have one loss-of-function TPMT allele leading to intermediate TPMT activity (heterozygous deficient or intermediate metabolizers). The TPMT*2, TPMT*3A, and TPMT*3C alleles account for about 95% of individuals with reduced levels of TPMT activity. NUDT15 deficiency is detected in <1% of patients of European or African ancestry. Among patients of East Asian ancestry (i.e., Chinese, Japanese, Vietnamese), 2% have two loss-of-function alleles of the NUDT15 gene, and approximately 21% have one loss-of-function allele. The p.R139C variant of NUDT15 (present on the *2 and *3 alleles) is the most commonly observed, but other less common loss-of-function NUDT15 alleles have been observed.

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Patients with thiopurine S-methyl transferase (TPMT) or nucleotide diphosphatase (NUDT15) deficiency may be at an increased risk of severe and life-threatening myelotoxicity if receiving conventional doses of azathioprine. Death associated with pancytopenia has been reported in patients with absent TPMT activity receiving azathioprine. In patients with severe myelosuppression, consider evaluation for TPMT and NUDT15 deficiency. Consider alternative therapy in patients with homozygous TPMT or NUDT15 deficiency and reduced dosages in patients with heterozygous deficiency.

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TPMT and NUDT15 Testing: Consider genotyping or phenotyping patients for TPMT deficiency and genotyping for NUDT15 deficiency in patients with severe myelosuppression. TPMT and NUDT15 testing cannot substitute for complete blood count (CBC) monitoring in patients receiving azathioprine. Accurate phenotyping (red blood cell TPMT activity) results are not possible in patients who have received recent blood transfusions.

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Patients with TPMT and/or NUDT15 Deficiency

Consider testing for TPMT and NUDT15 deficiency in patients who experience severe bone marrow toxicities. Early drug discontinuation may be considered in patients with abnormal CBC results that do not respond to dose reduction.

Homozygous deficiency in either TPMT or NUDT15 Because of the risk of increased toxicity, consider alternative therapies for patients who are known to have TPMT or NUDT15 deficiency.

Heterozygous deficiency in TPMT and/or NUDT15 Because of the risk of increased toxicity, dosage reduction is recommended in patients known to have heterozygous deficiency of TPMT or NUDT15. Patients who are heterozygous for both TPMT and NUDT15 deficiency may require more substantial dosage reductions.

Please review the complete therapeutic recommendations that are located here: (1).

TPMT recommendation

If starting doses are already high (e.g., 75 mg/m² of mercaptopurine), as is true in some ALL treatment regimens, lower than normal starting doses should be considered in TPMT intermediate metabolizers and markedly reduced doses (10-fold reduction) should be used in TPMT poor metabolizers. This approach has decreased the risk of acute toxicity without compromising relapse rate in ALL. Even at these markedly reduced dosages, erythrocyte TGN concentrations in TPMT poor metabolizers remain well above those tolerated and achieved by the majority of patients (who are TPMT normal metabolizers).

In some nonmalignant conditions, alternative agents may be chosen for TPMT intermediate or poor metabolizers rather than reduced doses of thiopurines; if thiopurines are used, full starting doses are recommended for TPMT normal metabolizers, reduced doses (30–80% of target dose) in TPMT intermediate metabolizers, and substantially reduced doses (or use of an alternative agent) in TPMT poor metabolizers.

Some of the clinical data upon which dosing recommendations are based rely on measures of TPMT phenotype rather than genotype; however, because TPMT genotype is strongly linked to TPMT phenotype, these recommendations apply regardless of the method used to assess TPMT status.

NUDT15 recommendation

Similar to TPMT, tolerated mercaptopurine dosage is also correlated with the number of nonfunctional alleles of the NUDT15 gene. In fact, the degree of thiopurine intolerance (e.g., for mercaptopurine) is largely comparable between carriers of TPMT vs. NUDT15 decreased function alleles, there remains a paucity of multi-ethnic studies examining both TPMT and NUDT15 variants.

Therefore, our NUDT15 recommendations parallel those for TPMT. For NUDT15 normal metabolizers (NUDT15*1/*1), starting doses do not need to be altered. For NUDT15 intermediate metabolizers (e.g., NUDT15*1/*3), reduced starting doses should be considered to minimize toxicity, particularly if the starting doses are high (e.g., 75 mg/m²/ day for mercaptopurine). For NUDT15 poor metabolizers (e.g., NUDT15*3/*3), substantially reduced doses (e.g., 10 mg/m²/ day of mercaptopurine) or the use of an alternative agent should be considered.

As for TPMT, there is substantial variability in the tolerated thiopurine dosages within NUDT15 intermediate metabolizers, with a minority of individuals who do not seem to require significant dose reduction. Therefore, genotype-guided prescribing recommendations apply primarily to starting doses; subsequent dosing adjustments should be made based on close monitoring of clinical myelosuppression (or disease-specific guidelines). In contrast, a full dose of mercaptopurine poses a severe risk of prolonged hematopoietic toxicity in NUDT15 poor metabolizers and pre-emptive dose reductions are strongly recommended.

The NUDT15 poor metabolizer phenotype is observed at a frequency of about 1 in every 50 patients of East Asian descent, which is more common than the TPMT poor metabolizer phenotype in Europeans, and, thus, genotyping NUDT15 in the Asian populations may be of particular clinical importance. NUDT15 deficiency is also more prevalent in individuals of Hispanic ethnicity, particularly those with high levels of Native American genetic ancestry.

Please review the complete therapeutic recommendations, which include CPIC’s recommended course of action if both TPMT and NUDT15 genotypes are known, located here: (2).

The Dutch Pharmacogenetics Working Group considers genotyping before starting azathioprine or 6-mercaptopurine to be essential for drug safety. Genotyping must be performed before drug therapy has been initiated to guide drug and dose selection.

TPMT Intermediate Metabolizer

Grade 2 leukopenia occurs in 23% of these patients with normal therapy for immunosuppression. The genetic variation increases the quantity of the active metabolites of azathioprine and mercaptopurine.

Recommendation:

IMMUNOSUPPRESSION

• Start with 50% of the standard dose

Adjustment of the initial dose should be guided by toxicity (monitoring of blood counts) and effectiveness.

Dose adjustment is not required for doses lower than 1.5 mg/kg per day for azathioprine or 0.75 mg/kg per day for mercaptopurine.

LEUKEMIA

• Start with 50% of the standard mercaptopurine dose, or start with the standard dose and reduce to 50% if side effects necessitate a dose reduction

It is not known whether dose reduction in advance results in the same efficacy as dose reduction based on toxicity.

The initial dose should be adjusted based on toxicity (monitoring of the blood counts) and efficacy.

Note: more stringent dose reductions are necessary if the patient is also NUDT15 IM or NUDT15 PM.

TPMT Poor Metabolizer

Grade 2 leukopenia and intolerance occurred in 98% of these patients with standard therapy. The gene variation increases the quantities of the active metabolites of azathioprine and mercaptopurine.

Recommendation:

• Choose an alternative or use 10% of the standard dose.

Any adjustment of the initial dose should be guided by toxicity (monitoring of blood counts) and effectiveness.

If the dose is decreased: advise patients to seek medical attention when symptoms of myelosuppression (such as severe sore throat in combination with fever, regular nosebleeds and tendency to bruising) occur

Background information:

Azathioprine is converted in the body to mercaptopurine. Mercaptopurine is an inactive pro-drug, which is converted to the active metabolites - thioguanine nucleotides - in the body.

Two catabolic routes reduce mercaptopurine bio-availability for thioguanine nucleotide formation. Thiopurine methyltransferase (TPMT) catalyses S-methylation of both mercaptopurine and the 6- mercaptopurine ribonucleotides formed in the metabolic pathway. In addition to this, mercaptopurine is oxidised to the inactive 6-thiouric acid by the enzyme xanthine oxidase (XO), which occurs primarily in the liver and intestines.

For more information about the TPMT phenotypes: see the general background information about TPMT on the KNMP Knowledge Bank or on www.knmp.nl (search for TPMT).

NUDT15 Intermediate Metabolizer

Grade ≥ 2 leukopenia occurs in 42% of these patients with standard immunosuppression therapy. The gene variation increases the concentration of the fully activated metabolite of azathioprine and mercaptopurine.

IMMUNOSUPPRESSION

• start with 50% of the standard dose

Adjustment of the initial dose should be performed based on toxicity (monitoring of the blood counts) and efficacy.

LEUKEMIA

• start at 50% of the standard mercaptopurine dose, or start with the standard dose and reduce to 50% if side effects necessitate a dose reduction

It is not known whether dose reduction in advance results in the same efficacy as dose reduction based on toxicity.

Adjustment of the initial dose should be performed based on toxicity (monitoring of the blood counts) and efficacy.

Note: more stringent dose reductions are necessary if the patient is also TPMT IM or TPMT PM.

NUDT15 Poor Metabolizer

Grade ≥ 2 leukopenia occurs in 96% of these patients with standard therapy. The gene variation increases the concentration of the fully activated metabolite of azathioprine and mercaptopurine.

• avoid azathioprine and mercaptopurine
• if it is not possible to avoid azathioprine and mercaptopurine: use 10% of the standard dose and advise patients to seek medical attention when symptoms of myelosuppression (such as severe sore throat in combination with fever, regular nosebleeds and tendency to bruising) occur

Any adjustment of the initial dose should be guided by toxicity (monitoring of blood counts) and efficacy.

Background information:

NUDT15 reverses the last step in the formation of the active metabolite of mercaptopurine and its precursor azathioprine. It converts 6-thiodeoxyguanosine triphosphate (6-thio-dGTP), which is incorporated in DNA, to 6-thiodeoxyguanosine monophosphate (6-thio-dGMP). Lower metabolic activity of NUDT15 therefore leads to increased intracellular concentrations of the active metabolite 6- thio-dGTP. This increases the risk of side effects, such as myelosuppression.

For more information about TPMT and NUDT15 phenotypes: see the general background information in the KNMP Knowledge Bank or on www.knmp.nl (search for TPMT or NUDT15).

Please review the complete therapeutic recommendations that are located here: (3, 4).